Quantum mechanical interpretation of nitrite reduction by cytochrome cd(1)nitrite reductase from Paracoccus pantotrophus

The reduction of nitrite to nitric oxide in respiratory denitrification is catalyzed by a cytochrome cd(1) nitrite reductase in Paracoccus pantotrophu s (formerly known as Thiosphaera pantotropha LMD 92.63). High-resolution st ructures are available for the fully oxidized [Fulop, V., Moir, J. W., Ferg uson, S. J., and Hajdu, J. (1995) Cell 81, 369-377; Baker, S. C., Saunders, N. F., Willis, A. C., Ferguson, S. J., Hajdu, J., and Fulop, V. (1997) J. Mol. Biol. 269, 440-455] and fully reduced forms of this enzyme, as well as for various intermediates in its catalytic cycle [Williams, P. A., Fulop, V., Garman, E. F., Saunders, N. F., Ferguson, S. J., and Hajdu, J. (1997) N ature 389, 406-412]. On the basis of these structures, quantum mechanical t echniques (QM), including density functional methods (DFT), were combined w ith simulated annealing (SA) and molecular mechanics techniques (MM) to cal culate the electronic distribution of molecular orbitals in the active site during catalysis. The results show likely trajectories for electrons, prot ons, substrates,and products in the process of nitrite reduction, and offer an interpretation of the reaction mechanism. The calculations indicate tha t the redox state of the d(1) heme and charges on two histidines in the act ive site orchestrate catalysis locally. Binding of nitrite to the reduced i ron is followed by proton transfer from His345 and His388 to one of the oxy gens of nitrite, creating a water molecule and an [Fe(II)-NO+] complex. Val ence isomerization within this complex gives [Fe(III)NO]. The release of NO from the ferric iron is influenced by the protonation state of His345 and IIis388, and by the orientation of NO on the d(1) heme. Return of Tyr25 to a hydrogen-bonding position between His345 and His388 facilitates product r elease, but a rebinding of Tyr25 to the oxidized iron may be bypassed in st eady-state catalysis.